Hypohidrotic ectodermal dysplasias (HED) are characterized by the triad of signs consisting of sparse hair, abnormal or missing teeth and inability to sweat. Recently, mutations in EDAR, a novel receptor of the Tumor Necrosis Factor Receptor Family (ThFR) were shown to be responsible for autosomal dominant and recessive forms of HED suggesting an essential role of this receptor in the process of ectodermal differentiation. However, the signaling mechanism(s) by which EDAR leads to ectodermal differentiation have not been defined. We have recently discovered that EDAR is capable of activating the NF-kappaB, JNK and cell death pathways. The overall goal of this proposal is further characterization of the above signaling activities of EDAR so as to better understand its role in ectodermal differentiation and in the pathogenesis of ectodermal dysplasias. This goal will be achieved through the following specific aims. In aim 1, deletion and site-directed mutagenesis of the EDAR cytoplasmic domain will be undertaken to map the domains and residues critical for NF-kappaB, JNK and cell death activation. The results of these studies will be also correlated with the clinical phenotype of ectodermal dysplasias associated with various known human and mouse EDAR mutations so as to explain the clinical heterogeneity of these disorders. In aim 2, mechanism(s) of EDAR-induced NF-kappaB, JNK and cell death pathways will be explored by using dominant negative inhibitors of the-proteins known to be involved in the activation of these pathways via other members of the TNFR family. Finally, in aim 3, yeast two- hybrid approach will be used to isolate novel signaling proteins that interact with the cytoplasmic domain of EDAR and their role in its various signaling activities explored. We hope that these studies will lead to a better understanding of the signaling pathways utilized by EDAR and of the processes involved in skin and hair differentiation. Ultimately, these studies will also lead to a better understanding of the pathogenesis of hypohidrotic ectodermal dysplasias caused by defect in EDAR-signaling.